Voltage-gated K? channels (Kv) and large-conductance, Ca2+-activated K? (BK) channels control the excitability of vascular smooth muscle. By controlling membrane potential, these channels indirectly regulate Ca 2+channel activity and hence Ca 2+ influx into these cells. Activation of Ca 2+channels causes a global, cellwide increase in intracellular Ca 2+that leads to vasoconstriction. In sharp contrast, we recently found that local, sub-cellular Ca 2+release events through ryanodine receptors (RyR) located in the sarcoplasmic reticulum ("Ca 2?sparks") indirectly relax vascular smooth muscle by activating BK channels and thereby hyperpolarizing smooth muscle cells. Membrane hyperpolarization causes smooth muscle relaxation because it decreases Ca 2?channel opening probability, which decreases intracellular Ca 2+.The experiments outlined in this proposal will investigate the cellular and molecular mechanisms controlling the function of Kv and BK channels in vascular smooth muscle. Our preliminary studies suggest that the Ca+-dependent phosphatase calcineurin modulates the function of Kv and BK channel function in vascular smooth muscle either directly, by controlling the phosphorylation state of these channels, or indirectly, through its control of the transcription factor nuclear factor of activated T cells (NFAT). Furthermore, recent data suggests that the molecular composition and function of BK channels is altered during hypertension. Over the next five years we plan to test four specific hypotheses. First, that activation of calcineurin modifies the communication between BK channels and ryanodine receptors in vascular smooth muscle. Second, that reduced expression of the 131subunit of BK channels during hypertension reduces the sensitivity of these channels to physiological changes in Ca 2?.Third, that activation of NFAT leads to changes in the expression of Kv and BK channels in cerebral vascular smooth muscle. Fourth, that local Ca 2+signals control the nuclear translocation of NFAT in vascular smooth muscle. These experiments will provide new fundamental information on the mechanisms controlling Kv and BK channel function in cerebral vascular smooth muscle and will provide insights into the mechanisms underlying vasospasm, stroke and hypertension.